The objective of the proposed study is to measure estrogen metabolites in urine samples from 250 healthy women before and after a low fat and cholesterol diet intervention and compare values with those from a control sample of 250 women. A high fat diet has been linked to excess risk of breast cancer. The underlying mechanisms for this relationship is unknown but is thought to involve an impact of dietary fat on estrogen metabolites are 2- and 16alpha-hydroxyestrone (16alpha-OHE1 may posses initiator and promoter activities in non-transformed mammary epithelial cells. 16alpha-hydroxylation is elevated in breast cancer patients, in women at high risk for breast cancer and in strains of mice susceptible to mammary tumors. The proposed study will determine whether a reduction in dietary fat intake results in a shift from 16alpha-OHE1 to 2- hydroxyestrone in the urine. Given that 16a-OHE1 is a potent estrogen, urinary levels could potentially serve as biomarkers for breast cancer risk and results of this study using a new assay on large population should help to elucidate the relationship of dietary fat to estrogen metabolism and breast cancer risk. The significance for research of the proposed study lies in its utilization of a newly developed noninvasive assay of estrogen metabolites in conjunction with a large carefully controlled randomized trial of dietary fat/cholesterol reduction in healthy normal women. The proposed study will provide an excellent basis on which to proceed with population-based studies on determinants of estrogen metabolism and biomarkers of breast cancer risk. Studies such as this also have a major public health significance in helping to evaluate general dietary recommendations directed at health promotion.